Excessive mobilization of body reserves during the transition from pregnancy to lactation imposes a risk for metabolic diseases on dairy cows. We aimed to establish an experimental model for high v. normal mobilization and herein characterized performance, metabolic and endocrine changes from 7 weeks antepartum (a.p.) to 12 weeks postpartum (p.p.). Fifteen weeks a.p., 38 pregnant multiparous Holstein cows were allocated to two groups that were fed differently to reach either high or normal body condition scores (HBCS: 7.2 NEL MJ/kg dry matter (DM); NBCS: 6.8 NEL MJ/kg DM) at dry-off. Allocation was also based on differences in body condition score (BCS) in the previous and the ongoing lactation that was further promoted by feeding to reach the targeted BCS and back fat thickness (BFT) at dry-off (HBCS: >3.75 and >1.4 cm; NBCS: <3.5 and <1.2 cm). Thereafter, both groups were fed identical diets. Blood samples were drawn weekly from 7 weeks a.p. to 12 weeks p.p. to assess the serum concentrations of metabolites and hormones. The HBCS cows had greater BCS, BFT and BW than the NBCS cows throughout the study and lost more than twice as much BFT during the first 7 weeks p.p. compared with NCBS. Milk yield and composition were not different between groups, except that lactose concentrations were greater in NBSC than in HBCS. Feed intake was also greater in NBCS, and NBCS also reached a positive energy balance earlier than HBCS. The greater reduction in body mass in HBCS was accompanied by greater concentrations of non-esterified fatty acids, and β-hydroxybutyrate in serum after calving than in NBCS, indicating increased lipomobilization and ketogenesis. The mean concentrations of insulin across all time-points were greater in HBCS than in NBCS. In both groups, insulin and IGF-1 concentrations were lower p.p than in a.p. Greater free thyroxine (fT4) concentrations and a lower free 3-3′-5-triiodothyronine (fT3)/fT4 ratio were observed in HBCS than in NBCS a.p., whereas p.p. fT3/fT4 ratio followed a reverse pattern. The variables indicative for oxidative status had characteristic time courses; group differences were limited to greater plasma ferric reducing ability values in NBSC. The results demonstrate that the combination of pre-selection according to BCS and differential feeding before dry-off to promote the difference was successful in obtaining cows that differ in the intensity of mobilizing body reserves. The HBCS cows were metabolically challenged due to intense mobilization of body fat, associated with reduced early lactation dry matter intake and compromised antioxidative capacity.